Prolonged survival of organ allografts in rats can be produced by prior exposure of the recipient to donor antigens and antibodies to these antigens, phenomena called enhancement. We can transfer a state of specific unresponsiveness by injecting naive syngeneic recipients with thymocytes from enhanced graft-bearing recipients. The suppression has been shown to primarily affect development of cytotoxic T lymphocytes (CTL). We now plan to fractionate the thymus population on the basis of physical and surface marker properties so that we are able to determine which cell or cells are active, and the mode of action. Definition of the rat MHC (RTI) has progressed to the point where a variety of haplotype and regional differences are available. We have antibodies, some of them monoclonal, for several differentiation markers of the rat (Ia-1, Ia-2, Ag-F, ART-1, Thy 1, anti-"helper"), and more will be assessed. The specificity of the cells to antigen or idiotype will be determined by selective absorption of cells. In vitro systems of MLR, CML, and CTL will be used to assess the mode of action on development or effector phases of cellular immunity. Subcellular membrane fragments (Ia ion) not only generate primary proliferative (MLR-like) and CTL responses in vitro, but a potent suppressor population. This system will provide a source a suppressor cells for comparison to the in vitro generated cells. Laser activated flow cytometry will also be used to sort cells on the basis of surface markers. Finally, a current technique for propagation of T lymphocytes in vitro will be adapted to suppressor-rich populations to provide larger numbers of cells for study and for in vivo transfer.